Work to be pursued involves (1) Attempts to define the smallest autonomous replicating fragment of DNA derivable from Col E1. (2) Utilization of that DNA to define the necessary nucleotide sequence responsible for autonomous replication. (3) Utilization of that DNA to define DNA-protein interactions involved in Col E1 replication. (4) A study of protein synthesized "in vivo" from the Col E1 DNA genome. (5) The construction of a highly efficient cloning vehicle from Col E1.